The present invention relates to a sensing apparatus that detects analytes with the aid of an enhanced field created by allowing light to strike a detection surface at a specified angle of incidence.
Known as a method that can be used in bio-measurement (measurement of reactions in biomolecules) and the like to detect (or measure) analytes with high sensitivity and great ease is fluorometry in which fluorescence from a fluorescent material that is excited by light at a specified wavelength to emit fluorescence (i.e., a fluorescence emitting material) is detected to thereby detect (or measure) the analytes.
If the analytes in fluorometry are a fluorescent material, a sample of interest that is assumed to contain the analytes is irradiated with exciting light at a specified wavelength and the resulting emission of fluorescence is detected to verify the presence of the analytes.
Even if the analytes in fluorometry are not a fluorescent material, a specifically binding material, or a material that specifically binds to the analytes is labeled with a fluorescent material and then bound to the analytes; subsequently, the same procedure as described above is performed to detect fluorescence (specifically, the fluorescence from the fluorescent material with which the specifically binding material that has bound to the analytes is labeled), whereby the presence of the analytes is verified.
It has been proposed that the sensitivity of analyte detection in fluorometry be increased by exciting the fluorescent material with the aid of an enhanced electric field that results from surface plasmon resonance on a metal film (see, for example, JP 2002-62255 A, JP 2001-21565 A, and JP 2002-257731 A).
In each of the methods described in those patent documents, analytes labeled with a fluorescent material are positioned in the neighborhood of a thin metal film and light is allowed to strike the boundary surface between the thin metal film and a prism (either a semicylindrical or triangular glass prism) at an angle that satisfies the plasmon resonance condition (plasmon resonance angle) to create an enhanced electric field on the thin metal film so that the analytes in the neighborhood of the thin metal film are excited strong enough to amplify the emission of fluorescence from the fluorescent material. This is a method of fluorescence detection utilizing the surface plasmon enhanced fluorescence (which is hereinafter sometimes abbreviated as SPF).
As described in JP 2001-21565 A, the electric field of surface plasmons is highly localized on the metal surface and attenuates exponentially with the distance from the metal surface, so fluorescently labeled antibodies (i.e., the fluorescent material) adsorbed onto the metal surface can be excited selectively and with high probability. As also described in JP 2001-21565 A, this SPF-based version of fluorescence detection ensures that the effect of any interfering material that is distant from the interface is suppressed to the smallest level, which also allows for precise detection of the analytes.
JP 2001-21565 A and JP 2002-257731 A also describe a rotating mechanism that adjusts the angle of the prism that bears the metal film; the angle of the prism is adjusted by this rotating mechanism so that the light issued from the light source is allowed to be launched into the prism at an optimum plasmon resonance angle.
The method of detecting analytes by making use of the surface plasmon enhancing effect is not limited to detecting the fluorescence excited by surface plasmons and another method that can be adopted is by detecting scattered light.
JP 10-78390 A describes a surface plasmon sensor comprising a prism, a metal film formed on a surface of the prism, a functional thin film that is provided on a surface of the metal film to trap analytes by an antigen-antibody reaction, and a flow cell that supplies a sample liquid as it makes contact with the functional thin film.
The operating principle of this surface plasmon sensor is such that the electric field of surface plasmons as excited by making use of the surface plasmon enhancing effect on the metal film is disturbed by the analytes present on the functional thin film to generate scattered light, which is detected to eventually detect the analytes. Thus, the method that detects scattered light rather than fluorescence can also be utilized to detect analytes.